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1.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-1008754

RESUMEN

This study investigated the acute toxicity of fermented Platycodonis Radix on mice and its effect on coughing in mice infected with Mycoplasma pneumoniae. The maximum dosage(MAD) was used in the acute toxicity experiment on mice to observe the signs of mice. After 14 days, dissection, blood biochemical examination, and pathological tissue section observation were conducted. In the pharmacological experiment of fermented Platycodonis Radix, 60 healthy BALB/c mice, 30 males and 30 females, were randomly divided into a blank group, a model group, a carbetapentane group(0.013 g·kg~(-1)·d~(-1)), and high-, medium-, and low-dose fermented Platycodonis Radix groups(5.2, 2.6, and 1.3 g·kg~(-1)·d~(-1)), with 10 mice in each group. Except for the blank group, the mice in the other five groups underwent model induction by intranasally instilling 20 μL of 1×10~6 CCU M. pneumoniae for 3 days, and the mice in each group were orally administered the corresponding drugs for 7 days. Cough induction experiment was conducted to observe and record the cough latency and total cough count within 3 min for each group. Hematoxylin-eosin(HE) staining and Masson staining were used to observe the pathological changes in lung tissues. Immunohistochemistry was performed to observe the protein expression of transient receptor potential A1(TRPA1), calcitonin gene-related peptide(CGRP), and substance P(SP) in the lung tissues of mice in each group. Real-time fluorescence-based quantitative polymerase chain reaction(qRT-PCR) was used to elucidate the changes in the mRNA levels of cough-related factors TRPA1, CGRP, and SP in mice treated with fermented Platycodonis Radix. No mice died in the acute toxicity experiment, and there were no changes in general behavior and major organ histopathological examinations. Compared with the blank group, there were no statistically significant differences in blood biochemical indexes. In the pharmacological experiment of fermented Platycodonis Radix, compared with the model group, the high-and medium-dose fermented Platycodonis Radix groups showed improved lung tissue structure of mice, with clear structure and regular tissue morphology. The qRT-PCR and immunohistochemical detection showed a decrease in the expression of TRPA1, CGRP, and SP in the fermented Platycodonis Radix groups. Fermented Platycodonis Radix can exert an inhibitory effect on cough by suppressing the expression of TRPA1, CGRP, and SP in lung tissues, thereby identifying the target of the drug.


Asunto(s)
Animales , Femenino , Masculino , Ratones , Péptido Relacionado con Gen de Calcitonina/análisis , Tos , Medicamentos Herbarios Chinos/química , Pulmón , Raíces de Plantas/química
2.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-936356

RESUMEN

OBJECTIVE@#To develop a convenient method for rapid purification of fresh Pheretima proteins and assess the inhibitory effect of these proteins against pulmonary fibrosis.@*METHODS@#The crude extract of fresh Pheretima was obtained by freeze-drying method and then purified by size exclusion chromatography. The composition of the purified proteins was analyzed by mass spectrometry. MRC-5 cells were treated with 5 ng/mL TGF-β1 alone (model group) or in combination with SB431542 (2 μmol/L) or the purified proteins (13.125 μg/mL), and the cytotoxicity of purified proteins and their inhibitory effects on cell proliferation were detected with CCK8 assay. Flow cytometry was used to detect the changes in cell apoptosis, and the cellular expressions of α-SMA, Vimentin, E-cadherin, collagen I, Smad2/3 and P-Smad2/3 were detected using RT-PCR and Western blotting. In the animal experiment, adult male C57BL/6 mice were subjected to intratracheal instillation of bleomycin followed by treatment with the purified proteins (5 mg/mL) for 21 days, after which HE and Masson staining was used to observe the pathological changes in the lung tissue of the mice.@*RESULTS@#We successfully obtained purified proteins from fresh Pheretima protein by size exclusion chromatography. Treatment with the purified proteins significantly inhibited TGF-β1-induced proliferation of MRC-5 cells (P < 0.01), reduced the cellular expressions of α-SMA, Vimentin and collagen I (P < 0.001 or P < 0.01), increased the expression of E-cadherin (P < 0.01), and inhibited the expressions of Smad2/3 and P-Smad2/3 (P < 0.001 or P < 0.01). In male C57BL/6 mice models of bleomycin-induced pulmonary fibrosis, treatment with the purified proteins obviously reduced the number of inflammatory cells and fibrotic area in the lungs.@*CONCLUSION@#The purified proteins from fresh Pheretima obtained by size exclusion chromatography can inhibit pulmonary fibrosis in mice by regulating the TGF-β/ Smad pathway.


Asunto(s)
Animales , Masculino , Ratones , Productos Biológicos/farmacología , Bleomicina/efectos adversos , Cadherinas/metabolismo , Colágeno Tipo I , Pulmón/patología , Ratones Endogámicos C57BL , Oligoquetos/química , Fibrosis Pulmonar/tratamiento farmacológico , Factor de Crecimiento Transformador beta1/metabolismo , Vimentina/metabolismo
3.
Nature ; 583(7818): 834-838, 2020 07.
Artículo en Inglés | MEDLINE | ID: mdl-32408338

RESUMEN

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel coronavirus with high nucleotide identity to SARS-CoV and to SARS-related coronaviruses that have been detected in horseshoe bats, has spread across the world and had a global effect on healthcare systems and economies1,2. A suitable small animal model is needed to support the development of vaccines and therapies. Here we report the pathogenesis and transmissibility of SARS-CoV-2 in golden (Syrian) hamsters (Mesocricetus auratus). Immunohistochemistry assay demonstrated the presence of viral antigens in nasal mucosa, bronchial epithelial cells and areas of lung consolidation on days 2 and 5 after inoculation with SARS-CoV-2, followed by rapid viral clearance and pneumocyte hyperplasia at 7 days after inoculation. We also found viral antigens in epithelial cells of the duodenum, and detected viral RNA in faeces. Notably, SARS-CoV-2 was transmitted efficiently from inoculated hamsters to naive hamsters by direct contact and via aerosols. Transmission via fomites in soiled cages was not as efficient. Although viral RNA was continuously detected in the nasal washes of inoculated hamsters for 14 days, the communicable period was short and correlated with the detection of infectious virus but not viral RNA. Inoculated and naturally infected hamsters showed apparent weight loss on days 6-7 post-inoculation or post-contact; all hamsters returned to their original weight within 14 days and developed neutralizing antibodies. Our results suggest that features associated with SARS-CoV-2 infection in golden hamsters resemble those found in humans with mild SARS-CoV-2 infections.


Asunto(s)
Betacoronavirus/patogenicidad , Infecciones por Coronavirus/transmisión , Infecciones por Coronavirus/virología , Modelos Animales de Enfermedad , Pulmón/patología , Pulmón/virología , Mesocricetus/virología , Neumonía Viral/transmisión , Neumonía Viral/virología , Aerosoles , Células Epiteliales Alveolares/patología , Células Epiteliales Alveolares/virología , Animales , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Antígenos Virales/inmunología , Antígenos Virales/aislamiento & purificación , Antígenos Virales/metabolismo , Betacoronavirus/inmunología , Betacoronavirus/aislamiento & purificación , Betacoronavirus/metabolismo , Bronquios/patología , Bronquios/virología , COVID-19 , Infecciones por Coronavirus/inmunología , Duodeno/virología , Fómites/virología , Vivienda para Animales , Riñón/virología , Masculino , Mesocricetus/inmunología , Mucosa Nasal/virología , Pandemias , Neumonía Viral/inmunología , ARN Viral/análisis , SARS-CoV-2 , Carga Viral , Pérdida de Peso
4.
mBio ; 11(2)2020 03 03.
Artículo en Inglés | MEDLINE | ID: mdl-32127444

RESUMEN

Anti-galactose-α-1,3-galactose (anti-α-Gal) antibody is naturally expressed at a high level in humans. It constitutes about 1% of immunoglobulins found in human blood. Here, we designed a live attenuated influenza virus vaccine that can generate α-Gal epitopes in infected cells in order to facilitate opsonization of infected cells, thereby enhancing vaccine-induced immune responses. In the presence of normal human sera, cells infected with this mutant can enhance phagocytosis of human macrophages and cytotoxicity of NK cells in vitro Using a knockout mouse strain that allows expression of anti-α-Gal antibody in vivo, we showed that this strategy can increase vaccine immunogenicity and the breadth of protection. This vaccine can induce 100% protection against a lethal heterosubtypic group 1 (H5) or group 2 (mouse-adapted H3) influenza virus challenge in the mouse model. In contrast, its heterosubtypic protective effect in wild-type or knockout mice that do not have anti-α-Gal antibody expression is only partial, demonstrating that the enhanced vaccine-induced protection requires anti-α-Gal antibody upon vaccination. Anti-α-Gal-expressing knockout mice immunized with this vaccine produce robust humoral and cell-mediated responses upon a lethal virus challenge. This vaccine can stimulate CD11blo/- pulmonary dendritic cells, which are known to be crucial for clearance of influenza virus. Our approach provides a novel strategy for developing next-generation influenza virus vaccines.IMPORTANCE Influenza A viruses have multiple HA subtypes that are antigenically diverse. Classical influenza virus vaccines are subtype specific, and they cannot induce satisfactory heterosubtypic immunity against multiple influenza virus subtypes. Here, we developed a live attenuated H1N1 influenza virus vaccine that allows the expression of α-Gal epitopes by infected cells. Anti-α-Gal antibody is naturally produced by humans. In the presence of this antibody, human cells infected with this experimental vaccine virus can enhance several antibody-mediated immune responses in vitro Importantly, mice expressing anti-α-Gal antibody in vivo can be fully protected by this H1N1 vaccine against a lethal H5 or H3 virus challenge. Our work demonstrates a new strategy for using a single influenza virus strain to induce broadly cross-reactive immune responses against different influenza virus subtypes.


Asunto(s)
Reacciones Cruzadas/inmunología , Epítopos/inmunología , Galactosa/inmunología , Inmunogenicidad Vacunal , Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Animales , Humanos , Virus de la Influenza A/clasificación , Virus de la Influenza A/inmunología , Vacunas contra la Influenza/genética , Células Asesinas Naturales/inmunología , Macrófagos/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Vacunas Atenuadas/genética , Vacunas Atenuadas/inmunología
6.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-878836

RESUMEN

To investigate the effect of baicalin extracted from Qinbai Qingfei Concentrated Pills on the expressions of TGF-β1, mmp2 and timp2 in mice with pulmonary fibrosis induced by bleomycin. The Biacore technique was used to detect the specific binding between Qinbai Qingfei Concentrated Pills and TGF-β1, and the affinity components were enriched, regenerated and recovered by Biacore fishing. Then ultra-performance liquid chromatography and quadrupole time of flight mass spectrometry(UPLC-Q-TOF-MS) were used to determine whether the monomer was baicalin. Biacore was used to verify the affinity kinetics of baicalin, which was validated by pharmacodynamics in vivo. Totally 30 BALB/C mice were randomly divided into three groups: baicalin group, blank group and model group. The blank group was given sodium chloride injection(0.08 mL·kg~(-1)), while the model group and the baicalin group were injected with 4 mg·kg~(-1) bleomycin. The localization of TGF-β1, mmp2 and timp2 protein in the cells and the mRNA expressions of TGF-β1, mmp2 and timp2 were detected by RT-PCR 14 days later. The results of Biacore affinity analysis showed that the peak of binding response between Qinbai Qingfei Concentrated Pills and TGF-β1 protein reached 1 524.0 RU, with specific binding. The affinity constant K_D of baicalin and TGF-β1 was 1.620 06 μmol·L~(-1), which was determined by SPR kinetic analysis, suggesting a stable binding between baicalin and TGF-β1, which verified the results of angulation. The results of immunohistochemistry showed that the deposition of cellulose in baicalin group was significantly less than that in model group, the mRNA expressions of TGF-β1, mmp2 and timp2 were decreased in baicalin solution compared with the model group. Baicalin combined with TGF-β1 could inhibit the expressions of mmp2 and timp2 and delay the progress of pulmonary fibrosis.


Asunto(s)
Animales , Ratones , Flavonoides , Cinética , Ratones Endogámicos BALB C , Fibrosis Pulmonar , Factor de Crecimiento Transformador beta1
7.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-1008423

RESUMEN

The aim of this paper was to investigate the effect of Dilong( geosaurus) on the expressions of fibrotic factors TGF-β1 and α-SMA in bleomycin-induced pulmonary fibrosis mice. The binding ability of Dilong to fibrotic factor TGF-β1 was initially detected by Biacore technology and verified by in vivo pharmacodynamics. A total of 60 SPF C57 mice were randomly divided into 6 groups. Except the blank group( injecting 0. 08 m L·kg-1 sodium chloride in the trachea),the other five groups were given bleomycin( 4 mg·kg-1) to replicate the pulmonary fibrosis model. After 14 days of drug treatment,the expressions of TGF-β1 and α-SMA were detected by Masson staining,immunohistochemistry and RT-PCR. The results of Biacore experiment showed that the extract of Dilong was well bound to TGF-β1 protein in vitro,and the binding value reached 619. 3. Compared with the model group,Masson's results showed that cellulose deposition in high-dose,medium-dose and low-dose Dilong groups decreased to varying degrees. RT-PCR results showed that different doses of Dilong could reduce protein and mRNA expressions of TGF-β1 and α-SMA to a certain extent in a dose-dependent manner. In conclusion,Dilong could delay the process of pulmonary fibrosis by binding to target protein TGF-β1 and inhibiting the expression of α-SMA.


Asunto(s)
Animales , Ratones , Actinas/metabolismo , Bleomicina , Pulmón , Medicina Tradicional China , Ratones Endogámicos C57BL , Oligoquetos , Fibrosis Pulmonar/metabolismo , Distribución Aleatoria , Factor de Crecimiento Transformador beta1/metabolismo
8.
Virology ; 525: 73-82, 2018 12.
Artículo en Inglés | MEDLINE | ID: mdl-30248524

RESUMEN

The limited protection of current commerical vaccines necessitates the investigation of novel vaccine strategies for unpredictable outbreaks. To investigate the feasibility of using vaccines derived from Group 1 influenza A virus to induce broadly cross-reactive immune responses against multiple influenza subtypes, we tested a panel of sequential 4-dose immunization regimens in mice. Mice were treated with inactivated (seasonal H1N1, pandemic H1N1 and H5N1) and vaccinia virus-based H5N1 live-attenuated vaccines in different combinations. Mice were then challenged by viruses of either Group 1 (H1N1) or Group 2 (H3N2, H7N7) influenza virus. All studied sequential 4-dose vaccinations could induce some degrees of heterosubtypic protection in mice. Amongst all these regimens, the combined use of inactivated and live-attenuated vaccines could achieve the best heterologous protection. These results highlight the synergistic effect of combining different vaccine platforms to enhance heterosubtypic protection against influenza viruses.


Asunto(s)
Vacunas contra la Influenza/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Animales , Antígenos Virales , Femenino , Vacunas contra la Influenza/administración & dosificación , Ratones , Ratones Endogámicos BALB C , Vacunas Atenuadas/administración & dosificación , Vacunas Atenuadas/inmunología , Vacunas de Productos Inactivados/administración & dosificación , Vacunas de Productos Inactivados/inmunología
9.
Front Immunol ; 9: 1479, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30013557

RESUMEN

Influenza viruses circulate worldwide causing annual epidemics that have a substantial impact on public health. This is despite vaccines being in use for over 70 years and currently being administered to around 500 million people each year. Improvements in vaccine design are needed to increase the strength, breadth, and duration of immunity against diverse strains that circulate during regular epidemics, occasional pandemics, and from animal reservoirs. Universal vaccine strategies that target more conserved regions of the virus, such as the hemagglutinin (HA)-stalk, or recruit other cellular responses, such as T cells and NK cells, have the potential to provide broader immunity. Many pre-pandemic vaccines in clinical development do not utilize new vaccine platforms but use "tried and true" recombinant HA protein or inactivated virus strategies despite substantial leaps in fundamental research on universal vaccines. Significant hurdles exist for universal vaccine development from bench to bedside, so that promising preclinical data is not yet translating to human clinical trials. Few studies have assessed immune correlates derived from asymptomatic influenza virus infections, due to the scale of a study required to identity these cases. The realization and implementation of a universal influenza vaccine requires identification and standardization of set points of protective immune correlates, and consideration of dosage schedule to maximize vaccine uptake.

10.
PLoS One ; 8(3): e58970, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23527059

RESUMEN

BACKGROUND: The radiation-induced energy metabolism dysfunction related to injury and radiation doses is largely elusive. The purpose of this study is to investigate the early response of energy metabolism in small intestinal tissue and its correlation with pathologic lesion after total body X-ray irradiation (TBI) in Tibet minipigs. METHODS AND RESULTS: 30 Tibet minipigs were assigned into 6 groups including 5 experimental groups and one control group with 6 animals each group. The minipigs in these experimental groups were subjected to a TBI of 2, 5, 8, 11, and 14 Gy, respectively. Small intestine tissues were collected at 24 h following X-ray exposure and analyzed by histology and high performance liquid chromatography (HPLC). DNA contents in this tissue were also examined. Irradiation causes pathologic lesions and mitochondrial abnormalities. The Deoxyribonucleic acid (DNA) content-corrected and uncorrected adenosine-triphosphate (ATP) and total adenine nucleotides (TAN) were significantly reduced in a dose-dependent manner by 2-8 Gy exposure, and no further reduction was observed over 8 Gy. CONCLUSION: TBI induced injury is highly dependent on the irradiation dosage in small intestine and inversely correlates with the energy metabolism, with its reduction potentially indicating the severity of injury.


Asunto(s)
Metabolismo Energético/efectos de la radiación , Intestino Delgado/metabolismo , Intestino Delgado/efectos de la radiación , Traumatismos por Radiación/metabolismo , Porcinos Enanos/metabolismo , Nucleótidos de Adenina/metabolismo , Animales , Daño del ADN/efectos de la radiación , Intestino Delgado/patología , Masculino , Mitocondrias/metabolismo , Mitocondrias/efectos de la radiación , Mitocondrias/ultraestructura , Dosis de Radiación , Porcinos , Factores de Tiempo , Triacetonamina-N-Oxil/metabolismo , Irradiación Corporal Total
11.
J Radiat Res ; 53(4): 537-44, 2012 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-22843618

RESUMEN

This study was undertaken to assess the diagnostic value of 2-[(18)F]-fluoro-2-deoxy-D-glucose positron emission tomography with computed tomography ([(18)F]-FDG-PET/CT) in the detection of radiation toxicity in normal bone marrow using Tibet minipigs as a model. Eighteen Tibet minipigs were caged in aseptic rooms and randomly divided into six groups. Five groups (n = 3/group) were irradiated with single doses of 2, 5, 8, 11 and 14 Gy of total body irradiation (TBI) using an 8-MV X-ray linear accelerator. These pigs were evaluated with [(18)F]-FDG-PET/CT, and their marrow nucleated cells were counted. The data were initially collected at 6, 24 and 72 h after treatment and were then collected on Days 5-60 post-TBI at 5-day intervals. At 24 and 72 h post-TBI, marrow standardized uptake value (SUV) data showed a dose-dependent decrease in the radiation dose range from 2-8 Gy. Upon long-term observation, SUV and marrow nucleated cell number in the 11-Gy and 14-Gy groups showed a continuous and marked reduction throughout the entire time course, while Kaplan-Meier curves of survival showed low survival. In contrast, the SUVs in the 2-, 5- and 8-Gy groups showed early transient increases followed by a decline from approximately 72 h through Days 5-15 and then normalized or maintained low levels through the endpoint; marrow nucleated cell number and survival curves showed approximately the same trend and higher survival, respectively. Our findings suggest that [(18)F]-FDG-PET/CT may be helpful in quickly assessing the absorbed doses and predicting the prognosis in patients.


Asunto(s)
Fluorodesoxiglucosa F18 , Sistema Hematopoyético/efectos de los fármacos , Tomografía de Emisión de Positrones/métodos , Tomografía Computarizada por Rayos X/métodos , Animales , Células de la Médula Ósea/citología , Supervivencia Celular , Relación Dosis-Respuesta en la Radiación , Masculino , Aceleradores de Partículas , Radiofármacos , Porcinos , Porcinos Enanos , Factores de Tiempo , Rayos X
12.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-237629

RESUMEN

This study was purposed to investigate the effects of TGF-beta1 and arsenic trioxide (As₂O₃) on cell apoptosis, cell cycle and changes of P27(Kip1), endogenous TGF-ß1, cyclin E and BCL-2 in HL-60 cells, and to explore the relationship between expression of P27(Kip1) and apoptosis induced by As₂O₃ and/or TGFß1. Cell apoptosis and cell cycle changes of HL-60 cells treated with As₂O₃ and/or TGFß1 were detected by cytomorphologic observation and flow cytometry, the protein expressions of P27(Kip1), TGF-ß1, cyclin E and BCL-2 were measured by immunohistochemistry. The results showed the effect of 5 μmol/L of As₂O₃ was the most strong among the different concentration of As₂O₃ ,and the effect on apoptosis at 48 hour was more strong than that at 24 hours (p < 0.05). The TGF-beta1 (5 ng/ml) induced arrest of cells in G₁ phase (p < 0.05) compared with As₂O₃ alone and As₂O₃ combined with TGF-ß1, while there was no difference with control. P27(Kip1) expression was up regulated (p < 0.05), cyclin E and BCL-2 expression was down-regulated (p < 0.05) in TGFß1-treated group. BCl-2 expression was down regulated, endogenesis TGFß1 expression was up regulated (p < 0.05), and the level of P27(kip1) and cyclin E were not changed in As₂O₃-treated group (p > 0.05). The down-regulating effect of TGF-ß1 combined with As₂O₃ on BCL-2 protein was more strong than that in single factor treated group (p < 0.05). It is concluded that TGFß1 induces cell cycle arrest and apoptosis in HL-60 cells, while the P27(kip1) expression is up regulated. P27 protein is the key effector of TGFß-induced cell cycle arrest. The effect of TGF-ß1 combined with As₂O₃ on apoptosis as well as the down-regulation of BCL-2 protein in HL-60 cells is more strong than that in single factor-treated groups, that indicates the passages linking up each other.


Asunto(s)
Humanos , Apoptosis , Arsenicales , Farmacología , Ciclina E , Metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , Metabolismo , Regulación Leucémica de la Expresión Génica , Células HL-60 , Óxidos , Farmacología , Proteínas Proto-Oncogénicas c-bcl-2 , Metabolismo , Factor de Crecimiento Transformador beta1 , Farmacología
13.
Chinese Journal of Biotechnology ; (12): 111-114, 2004.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-305218

RESUMEN

The biosensor based on optical imaging ellipsometry, can be used to detect directly, without labeling, the surface concentration of biomolecules on solid surface. The feasibility of using protein A to immobilize antibody on the silicon surface of the imaging ellipsometry biosensor was investigated in this study. The results showed that the anti-IgG immobilized by the protein A on silicon surface could bind effectively human IgG, and the human IgG immobilized on silicon surface by protein A bound more polyclonal antibody molecules than that immobilized on silicon surface directly, suggesting that protein A might block the surface to prevent the absorption of human IgG on surface directly, which might compromise its native configuration. The silicon surface modified with protein A is expected to be used to immobilize a variety of antibodies, as protein A can bind selectively the Fc regions of many mammalian IgG. The combination of imaging ellipsometry and the protein A surface modification has the potential to be developed into immunoassays of high sensitivity.


Asunto(s)
Humanos , Técnicas Biosensibles , Métodos , Inmunoensayo , Métodos , Inmunoglobulina G , Química , Proteína Estafilocócica A , Química
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